The aim of this proposal is to develop a new class of anthrax vaccines that can be produced rapidly and mass-administered by non-medical personnel during an emergency. It has been documented that intranasal inoculation of replication-defective adenovirus (Ad)-vectored vaccines is effective and safe. Unlike gene therapy, pre-existing immunity to Ad does not interfere with the potency of an Ad-vectored nasal vaccine. This class of vaccines can be produced quickly in response to an unpredicted escalation in demand. Nasal spray of an Ad mist can be performed by personnel with a low level of skill. These studies will determine whether Ad vectors encoding Bacillus anthracis protective antigen (PA83) and an atoxic fragment of immune inhibitor A (lnhAmut) can mobilize the immune repertoire toward a beneficial immune protection against inhalational anthrax. In this project, the potency of an Ad vector encoding PA83 will be compared to that of a codon-optimized counterpart after intranasal administration into mice and rabbits. An Ad vector encoding lnhAmut will be investigated for its potential to arrest spore germination by eliciting an immune response that neutralizes secretory immune inhibitor A. Whether lnhAmut can be formulated with PA83 as an adjunct anthrax vaccine will be determined. The fate of Ad vectors after intranasal administration and the safety profile will be analyzed. Protection of immunized rabbits against spores of the Ames isolate of B. anthracis will be evaluated. Once solid protection against anthrax spores is demonstrated in animals, clinical lots of Ad vectors will be produced under GMP (Good Manufacturing Practice) in anticipation of a future Phase I human trial. The overall goal of these milestones is to provide an easy-to-administer anthrax vaccine that fortifies civilians and military personnel within targeted regions against mass exposure to anthrax spores during a terrorist attack.